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Journal of Ethnic Foods
journal homepage: http://journalofethnicfoods.net
Original article
Antiobesity effects of kimchi in diet-induced obese mice
Meizi Cui a, Hee-Young Kim a, Kyung Hee Lee a, Ji-Kang Jeong a'b, Ji-Hee Hwang c, Kyu-Young Yeo c, Byung-Hee Ryu c, Jung-Ho Choi c, Kun-Young Park a'b' *
a Department of Food Science and Nutrition, Pusan National University, Busan, South Korea b Kimchi Research Institute, Pusan National University, Busan, South Korea c Daesang FNF Corporation R&D Center, Icheon, Gyongki, South Korea
ARTICLE INFO ABSTRACT
Background: The present study was investigated to confirm the antiobesity effect of kimchi in high-fat diet-induced obese C57BL/6 mice.
Methods: Mice in the high-fat diet (HFD) group, standardized kimchi (S-Kimchi) group, and Korean commercial kimchi (D-Kimchi) group, but not in the normal-diet group, were fed a high-fat diet (HFD) for the first 4 weeks to induce obesity. From the 5th to 8th weeks, the S- and D-Kimchi groups were fed an HFD containing 10% of S-Kimchi or D-Kimchi, respectively. After 8 weeks, mice were sacrificed and obesity-related factors were determined.
Results: Body and adipose tissue weights were significantly lower in the kimchi-treated groups than in the HFD group. In particular, in the D-Kimchi group, serum levels of triglyceride, total cholesterol, low-density lipoprotein-cholesterol, insulin, and leptin were significantly lower, and serum levels of high-density lipoprotein-cholesterol and adiponectin were markedly higher than those in the HFD group. Moreover, hepatic mRNA expression of adipogenesis-related genes (CCAAT/enhance-binding protein-a, peroxisome proliferator-activated receptor-g, sterol regulatory element-binding protein-lc, and fatty acid synthase) in the kimchi-treated groups were lower than those in the HFD group, but fatty acid oxidation-related carnitine palmitoyltransferase-1 expressions were higher. In addition, kimchi decreased the mRNA levels of the inflammation-related monocyte chemotactic protein-1 and interleukin-6 in epididymal fat tissue.
Conclusion: Administration of kimchi, especially of D-Kimchi, which contained Leuconostoc mesenteroides DRC 0211 starter and other subingredients, exhibited antiobesity activity by reducing body weight gains and adipose tissue weights; modulating serum lipid profiles and hepatic lipogenesis; regulating serum insulin, leptin, and adiponectin levels; and reducing adipocyte size and inflammatory response in epididymal fat tissues.
Copyright © 2015, Korea Food Research Institute, Published by Elsevier. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
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Article history: Received 17 May 2015 Received in revised form 27 June 2015 Accepted 8 July 2015 Available online 28 August 2015
Keywords: antiobesity C57BL/6 mice high-fat diet kimchi
obesity-related genes
1. Introduction
Kimchi is a traditional Korean fermented vegetable food prepared with baechu cabbage and other beneficial subingredients by lactic acid bacterial fermentation at a low temperature (4°C) [1]. Kimchi is low in calories (~18 kcal/100 g) and contains high levels of vitamins (such as vitamins A, C, B complex, K, and others), b-carotene, minerals (such as Na, Ca, K, Fe, and P), dietary fiber, and
* Corresponding author. Department of Food Science and Nutrition, Pusan National University, 2 Busandaehak-ro, 63-bunkil, Geumjeong-gu, Busan 609-735, South Korea.
E-mail address: kunypark@pusan.ac.kr (K.-Y. Park).
functional phytochemicals (such as benzyl isothiocyanate, indoles, thiocyanates, and sitosterols) that promote human health [1].
Many authors have reported that kimchi has beneficial properties, which include antimutagenic [2], anticancer [3], antioxidant [4], antiaging [5], antiatherogenic [6], and antidiabetic [7] activities, and fibrinolytic effects [8]. Furthermore, many studies have demonstrated the antiobesity effect of kimchi in animals [9,10] and humans [11]. Recently, it was reported that subingredients can improve the health benefits of kimchi [12,13], and that the inoculation of a pure culture starter, with Weissella koreensis or Leuco-nostoc (Leu.) citreum, increases the antiobesity effects of kimchi in high-fat diet-induced obese (DIO) mice [9], and improves the shelf life and quality of kimchi [14].
http://dx.doi.org/10.1016/j.jef.2015.08.001
2352-6181/Copyright © 2015, Korea Food Research Institute, Published by Elsevier. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/ licenses/by-nc-nd/4.0/).
Over recent years, the consumer trend is toward commercial kimchi. According to one report [15], over the past 7 years, the amount of kimchi consumed in Korea has remained constant, but the percentage accounted for by home-made kimchi has decreased (64.6% in 2007 and 60.0% in 2013), whereas commercial kimchi consumption has increased (35.3% in 2007 and 40.0% in 2013). Owing to the expanding commercial market, studies on kimchi-related probiotics and starter cultures have resulted in the development of commercial kimchis with added vegetables and in the use of lactic acid bacteria strains as a starter, such as Leu. meseteroides, Leu. citreum, or Lactobacillus plantarum, with constant quality, enhanced tastes, and health-promoting benefits.
Obesity, which is a component of metabolic syndrome, is associated with excess energy intake and low energy expenditure, and is characterized by weight gain, type 2 diabetes, hypertension, nonalcoholic fatty liver, glucose intolerance, inflammatory response, and coronary heart disease and cancer development associated with insulin resistance [16]. Obesity rates continue to increase worldwide and now exceed 30% in Mexico, New Zealand, and the United States. Thus, obesity is a major health problem worldwide [17].
The basic molecular imbalance driving obesity is that between lipogenesis and lipolysis. These processes are regulated in the liver and adipose tissues by multienzyme systems, such as CCAAT/ enhance-binding protein-a (C/EBP-a), sterol regulatory element-binding protein-lc (SREBP-lc), peroxisome proliferator-activated receptor-g (PPAR-g), fatty acid synthase (FAS), and carnitine palmitoyltransferase-1 (CPT-1) [18,19].
Some drugs used to treat clinical obesity are associated with adverse effects such as nausea, insomnia, constipation, gastrointestinal problems, and potential adverse cardiovascular effects [20]. Therefore, efforts are being made to find and develop antiobesity foods and food ingredients that effectively reduce body fat accumulation, reduce the risk of obesity-related chronic diseases, and minimize the side effects in clinical treatment [21,22].
The present study was designed to investigate the antiobesity activities of home-made and commercial kimchi (containing Leu. mesenteroides DRC 0211 as starter and several subingredients) in high-fat DIO mice, by determining weight loss effects after 4 weeks of obesity induction, followed by 4 weeks of kimchi administration. In addition, mechanisms responsible for their antiobesity effects were also investigated.
2. Materials and methods
2.1. Sample preparation
Standardized kimchi (S-Kimchi) was developed by the Kimchi Research Institute at Pusan National University (Busan, Korea), and prepared using the following ingredients: 13.0 g radish, 2.0 g green onion, 3.5 g red pepper powder, 1.4 g garlic, 0.6 g ginger, 2.2 g anchovy juice, and 1.0 g sugar, in 100 g of brined baechu cabbage [23].
A Korean commercial kimchi manufactured by Daesang FNF (D-Kimchi; Icheon, Korea), which was the most consumed commercial kimchi in 2012 [24], was used in the present study. D-Kimchi is prepared using the following ingredients: 18.0 g radish, 3.5 g red pepper powder, 3.8 g garlic, 0.5 g ginger, 2.5 g leek (Allium tuberosum), 1.5 g green onion, 3.5 g anchovy juice, 3 g sea tangle extract, and 8 g rice starch in 100 g of brined baechu cabbage; it was inoculated with 106 colony-forming units/g Leu. mesenteroides DRC 0211.
Kimchi samples were fermented at 4°C until pH values reached 4.3 (optimum for the ripened state) and then freeze-dried to prepare the experimental diets.
2.2. Animals and treatment
Male C57BL/6J mice (6-week old, 18 ± 0.5 g) were purchased from Samtako Bio Korea (Osan, Korea) and housed under standard conditions (50—60% relative humidity, under a 12-hour dark/12-hour light cycle). Food and water were supplied ad libitum.
Animals were randomly divided into four groups of eight mice each: the ND group was administered a normal diet (ND; AIN-93G diet) for 8 weeks; the HFD group was provided a high-fat diet (HFD; 45% lard oil in AIN-93G diet) for 8 weeks, and the S- and D-Kimchi groups were fed the HFD diet for the first 4 weeks and the HFD diet containing 10% of freeze-dried S-Kimchi or D-Kimchi, respectively, for the remaining 4 weeks. Compositions of the experimental diets are provided in Table 1.
Body weights and food intakes were checked weekly, and animals were sacrificed on the last day of the 8-week experimental period. The protocol used in this study was approved by the Institutional Animal Care and Use Committee of Pusan National University (PNU-IACUC) (approval number PNU-2013-0455).
2.3. Serum and tissue preparation
At the end of the experiment, blood samples were collected from the inferior vena cava of each animal. Serum was separated from blood by centrifugation at 3,000g for 10 minutes at 4°C. Organs of interest, that is, liver, epididymal fat, and perirenal fat tissues, were surgically removed immediately, washed with cold normal saline, wiped with a paper towel, and weighed. Serum and organ samples were stored at -80° C for further study.
2.4. Serum lipid, insulin, leptin, and adiponectin analysis
Serum levels of triglyceride (TG), total cholesterol (TC), high-density lipoprotein-cholesterol (HDL-c), and low-density lipopro-tein-cholesterol (LDL-c) were measured using a commercial kit (Asan Pharmaceutical Co., Seoul, Korea), and serum levels of insulin, leptin, and adiponectin were measured using a commercial enzyme-linked immunosorbent assay kit (R&D System, Minneapolis, MN, USA).
Table 1
Compositions of experimental diets used in the animal study.
Ingredient (g/1,000 g diet) Experimental diet
HFD S-Kimchi D-Kimchi
Casein 245 220.5 220.5
L-Cystine 3.5 3.15 3.15
Corn starch 85 76.5 76.5
Maltodextrin 115 103.5 103.5
Sucrose 200 180 180
Cellulose 58 52.2 52.2
Soybean oil 30 27 27
Lard 195 175.5 175.5
Mineral mix, A1N-93G-MX 43 38.7 38.7
Calcium phosphate, dibasic 3.4 3.06 3.06
Vitamin mix, A1N-93-VX 19 17.1 17.1
Choline bitartrate 3 2.7 2.7
Red food color 0.1 0.09 0.09
S-Kimchi — 100 —
D-Kimchi — — 100
Calories (kcal) 4,659 4,659 4,659
The ND group received ND (AIN-93G diet), the HFD group received a high-fat diet for 8 weeks, the S-Kimchi group received an HFD for 4 weeks followed by the same HFD containing 10% standard kimchi for 4 weeks, and the D-Kimchi group received the HFD for 4 weeks and HFD containing 10% commercial D-Kimchi for the next 4 weeks.
D-Kimchi, Korean commercial kimchi; HFD, high-fat diet; ND, normal diet; S-Kimchi, standardized kimchi.
2.5. Histological observations
The liver and epididymal fat tissues were subjected to histological examinations. Tissues were fixed in 10% (v/v) neutral buffered formalin, dehydrated in ethanol, and embedded in paraffin. Sections (4 mm) were then prepared and stained with hematoxylin and eosin. Images were developed using a Zeiss Axioskop 2 Plus microscope equipped with an AxioCam MRc5 CCD Camera (Carl Zeiss Microimaging, Thornwood, NY, USA).
2.6. Reverse transcription polymerase chain reaction assays
The mRNA expressions of C/EBP-a, SREBP-lc, PPAR-g, CPT-1, FAS, monocyte chemotactic protein-1 (MCP-1), and interleukin-6 (IL-6) were determined by reverse transcription polymerase chain reaction (PCR). TRIzol, oligo dT12-18 primer, reverse transcriptase buffer, deoxyribonucleotide triphosphates (dNTPs), murine Malo-ney leukemia virus reverse transcriptase, RNase inhibitor, and agarose were obtained from Invitrogen Life Technologies (Carlsbad, CA, USA). All were of analytical grade.
Total RNA was isolated using TRIzol reagent, according to the manufacturer's instructions. Briefly, 200 mL of chloroform was added to the liquid phase separated from tissue sample and centrifuged at 12,000g for 15 minutes at 25°C. Isopropanol was then added to supernatants at a 1:1 (v:v) ratio and RNA was pelleted by centrifugation (12,000g for 15 minutes at 4°C). After washing the pellets with 70% ethanol, the RNA was solubilized in diethyl pyrocarbonate-treated RNase-free water and quantified by measuring absorbance at 260 nm using a UV-2401PC spectrophotometer (Shimadzu, Kyoto, Japan). Equal amounts of RNA (1 mg) were then reverse transcribed in an AccuPower PCR PreMix (Bio-neer, Daejeon, Korea) containing 1 x reverse transcriptase buffer, 1mM dNTPs, 500 ng of oligo dT18 primers, 140 U of murine Maloney leukemia virus reverse transcriptase, and 40 U of RNase inhibitor for 45 minutes at 42° C. PCR was then carried out in an automatic PCR thermocycler (Bioneer). PCR products obtained were separated in 2% agarose gels and visualized by ethidium bromide staining. Glyceraldehyde-3-phosphate dehydrogenase was used for normalization.
2.7. Statistical analysis
Results are presented as means ± standard deviations. The significances of differences between group mean values were determined by one-way analysis of variance and Duncan's multiple range test. A value of p < 0.05 was considered statistically significant. SPSS v18.0 statistical software (SPSS Institute Inc., Chicago, IL, USA) was used to conduct the analysis.
3. Results
3.1. Effects of kimchis on body weights, food intakes, food efficiency ratios, and tissue weights
Initial body weights in the four study groups were similar. However, after the first four experimental weeks, the mean body weights in groups fed HFD, that is, the HFD, S-Kimchi, and D-Kimchi groups, were markedly higher than that of the ND group (29.5 ± 1.1 g, 29.1 ± 0.7 g, and 29.3 ± 0.7 g, respectively, vs. 24.7 ± 0.9 g) (Table 2). After the 8th week, the S- and D-Kimchi groups had significantly lower mean body weights than the HFD group (32.8 ± 2.5 g and 32.6 ± 2.3 g, respectively, vs. 43.0 ± 3.4 g) (p < 0.05). At this time, the ND group had a mean body weight of 27.2 ± 1.2 g. Summarizing, the mean body weight in the HFD group increased by 14 g after the 8-week experimental period, whereas the S- and D-Kimchi groups gained only 3 g, and the ND group gained 2.5 g.
No significant intergroup differences in food intake were observed; however, the mean food efficiency ratios in the S-Kimchi (3.9 ± 0.1%) and D-Kimchi (3.5 ± 0.5%) groups were significantly lower than that in the HFD group (13.9 ± 1.6%) (the mean food efficiency ratio in the ND group was 2.7 ± 0.8%) (Table 2).
After the 8-week experimental period, the mean liver weights in the kimchi groups (1.5 ± 0.2 g and 1.5 ± 0.1 g for the S- and D-Kimchi groups, respectively) were significantly lower than that in the HFD group (1.9 ± 0.5 g) (the mean liver weight in the ND group was 1.6 ± 0.1 g). Kimchi-treated groups showed significantly lower mean epididymal fat tissue weights (S-Kimchi group: 0.9 ± 0.4 g; D-Kimchi group: 0.9 ± 0.6 g) and perirenal fat tissue weights (S-Kimchi group: 0.5 ± 0.2 g; D-Kimchi group: 0.5 ± 0.3 g) than those in the HFD group (epididymal fat: 2.3 ± 0.5 g; perirenal fat: 1.0 ± 0.3 g) (the mean epididymal and perirenal fat tissue weights in the ND group were 0.4 ± 0.1 g and 0.1 ± 0.1 g, respectively; Fig. 1).
3.2. Effect of kimchi on serum lipid profiles
After the 8-week experimental period, HFD significantly increased serum levels of TG and TC in the HFD and S- and D-Kimchi groups as compared with the ND group (Table 3). The ND group (TG: 72.6 ± 4.7 mg/dL; TC: 85.1 ± 3.0 mg/dL) had the lowest TG and TC levels, and the HFD group had the highest levels (TG: 161.1 ± 10.9 mg/dL; TC: 124.1 ± 6.2 mg/dL). Serum levels of these markers in the S- and D-Kimchi groups were significantly lower than those in the HFD group, and the difference was greater for the D-Kimchi group (87.6 ± 8.4 mg/dL of TG and 90.0 ± 6.7 mg/dL of TC for the D-Kimchi group vs. 94.1 ± 9.3 mg/dL of TG and 104.4 ± 4.2 mg/dL of TC for the S-Kimchi group).
Table 2
Body weights, food intakes, food efficiency ratios, and tissue weights in DIO mice.
ND HFD S-Kimchi D-Kimchi
Body weight (g) Initial 23.4 ± 1.3 (NS) 23.3 t 1.3 23.5 10.6 23.4 t 0.6
4th wk 24.7 ± 0.9* (1.3)y 29.5 t 1.1z (6.2) 29.1 t 0.7z (5.6) 29.3 t 0.7z (5.9)
8th wk 27.2 ± 1.2x (2.5)" 43.0 t 3.4z (13.5) 32.8 t 2.5* (3.7) 32.6 t 2.3* (3.3)
Body weight gain (g/d) 0.1 ± 0.1* 0.5 t 0.1z 0.1 t 0.1* 0.1 t 0.1*
Food intake (g/d) 3.4 ± 0.1 (NS) 3.5 t 0.2 3.4 t 0.1 3.5 t 0.1
Food efficiency ratio (%)1 2.7 ± 0.8* 13.9 t 1.7z 3.9 t 0.1* 3.5 t 0.5*
Group definitions are detailed in the legend of Table 1. Results are presented as means ± SDs.
DIO, diet-induced obese; D-Kimchi, Korean commercial kimchi; HFD, high-fat diet; ND, normal diet; NS, not significant; SD, standard deviation; S-Kimchi, standardized kimchi. »4,5 Means with different letters in the same row are significantly different (p < 0.05) by Duncan's multiple range tests. y Grams of body weight gained during the first 4 weeks of the experimental period. 11 Grams of body weight gained during the second 4 weeks (after the 8-week experimental period). 1 Food efficiency ratio = total weight gain/total food intake.
Fig. 1. Tissue weights of liver, epididymal fat, and perirenal fat in DIO mice (after the 8-week experimental period). Results are presented as means ± SDs. Group definitions are detailed in the legend of Table 1. *,t,z Means with different letters are significantly different (p < 0.05) by Duncan's multiple range tests. DIO, diet-induced obese; D-Kimchi, Korean commercial kimchi; HFD, high-fat diet; ND, normal diet; SD, standard deviation; S-Kimchi, standardized kimchi.
Table 3
Serum levels of TG, TC, HDL-c, and LDL-c in DIO mice (after the 8-week experimental period).
Serum levels (mg/dL) Groups
ND HFD S-Kimchi D-Kimchi
TG 72.6 ± 4.7* 161.1 - t 10.9y 94.1 z H 9.3* 87.6 ± 8.4*,*
TC 85.1 ± 3.0x 124.1 - h 6.2y 104.4 - F 4.2« 90.0 ± 6.7*,x
LDL-c 9.8 ± 2.3* 50.8 - n6.0y 39.7 - .-1.9* 17.3 ± 6.8*
HDL-c 60.8 ± 2.2y 41.4 - t3.3* 45.8 z h 4.2* 55.2 ± 2.1y
Results are presented as means ± SDs. Group definitions are detailed in the legend of Table 1.
*,t,z Means with different letters in the same row are significantly different (p < 0.05) by Duncan's multiple range tests.
DIO, diet-induced obese; D-Kimchi, Korean commercial kimchi; HDL-c, high-density lipoprotein-cholesterol; HFD, high-fat diet; LDL-c, low-density lipoprotein-cholesterol; ND, normal diet; SD, standard deviation; S-Kimchi, standardized kimchi; TC, total cholesterol; TG, triglyceride.
At 8 weeks, the mean LDL-c level was greatest in the HFD group (50.8 ± 6.0 mg/dL), followed by the S-Kimchi group (39.7 ± 1.9 mg/ dL), D-Kimchi group (17.3 ± 6.8 mg/dL), and ND group (9.8 ± 2.3 mg/dL); the mean HDL-c level was low in the HFD group (41.4 ± 3.3 mg/dL), but significantly higher in the D-Kimchi group (55.2 ± 2.1 mg/dL) and higher in the S-Kimchi group (45.8 ± 4.2 mg/ dL). At this time, the ND group had a mean LDL-c level of 60.8 ± 2.2 mg/dL.
3.3. Effect of kimchi on serum levels of insulin, leptin, and adiponectin
At 8 weeks, the mean serum insulin level in the HFD group was significantly elevated (9.5 ± 0.1 mg/dL) as compared with that in
the S-Kimchi (7.2 ± 0.2 mg/dL), D-Kimchi (6.7 ± 0.4 mg/dL), and ND (6.1 ± 0.0 mg/dL) groups (p < 0.05).
The HFD group had the highest mean serum leptin level (39.2 ± 1.4 mg/dL) after the experimental period. Values for the other three groups were significantly lower (S-Kimchi group: 21.7 ± 1.0 mg/dL; D-Kimchi group: 17.2 ± 0.7 mg/dL; and ND group: 13.3 ± 0.9 mg/dL).
In addition, the mean adiponectin level was significantly higher in the D-Kimchi group (33.3 ± 0.2 mg/dL) than that in the S-Kimchi (31.7 ± 0.5 mg/dL) and HFD (29.0 ± 0.7 mg/dL) groups (the ND group had a mean adiponectin level of 35.4 ± 0.6 mg/dL; Fig. 2).
3.4. Histological observations and mRNA expression of C/EBP-a, PPAR-g, SREBP-1c, FAS, and CPT-1 in liver tissue
The hepatic histological study conducted after the 8-week experimental period showed that kimchi treatment significantly reduced HFD-induced lipid formation in the liver (Fig. 3A). The livers in the HFD group showed steatosis, characterized by numerous large fat globules. However, in the kimchi-treated groups, fat globules were obviously fewer and smaller. In particular, the livers in the D-Kimchi group appeared near normal; no fat globules were observed in the ND group.
Reverse transcription PCR was used to assess hepatic mRNA expressions of adipogenesis (C/EBP-a, PPAR-g, SREBP-1c, and FAS)-and fatty acid oxidation (CPT-1)-related genes. Interestingly, the D-Kimchi group showed lower expressions of adipogenesis-related factors than the S-Kimchi and HFD groups. In addition, kimchi supplementation also increased the mRNA levels of CPT-1 (a rate-limiting enzyme in the b-oxidation of fatty acids) as compared with that in the HFD group (Fig. 3B).
Fig. 2. Serum levels of insulin, adiponectin, and leptin in DIO mice after the 8-week experimental period. Results are presented as means ± SDs. Group definitions are detailed in the legend of Table 1. *,t,z,x Means with different letters are significantly different (p < 0.05) by Duncan's multiple range tests. DIO, diet-induced obese; D-Kimchi, Korean commercial kimchi; HFD, high-fat diet; ND, normal diet; SD, standard deviation; S-Kimchi, standardized kimchi.
Fig. 3. Study of the liver tissues of DIO mice after the 8-week experimental period. (A) Histological observations. (B) The mRNA expressions of C/EBP-a, PPAR-y, SREBP-1c, FAS, and CPT-1. The intensities of bands were measured by densitometry and are expressed as folds of the control (HFD) divided by GAPDH. Fold ratio: gene expression/GAPDH x control numerical value (control fold ratio set at 1). Group definitions are detailed in the legend of Table 1. *,t,M Means with different letters are significantly different (p < 0.05) by Duncan's multiple range tests. C/EBP-a, CCAAT/enhance-binding protein-a; CPT-1, carnitine palmitoyltransferase-1; DIO, diet-induced obese; D-Kimchi, Korean commercial kimchi; FAS, fatty acid synthase; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HFD, high-fat diet; ND, normal diet; PPAR-g, peroxisome proliferator-activated receptor-g; S-Kimchi, standardized kimchi; SREBP-1c, sterol regulatory element-binding protein-1c.
3.5. Histological observations and adipocytokines in epididymal fat tissues
Histological assays showed that kimchi administration reduced HFD-induced fat accumulation in epididymal fat tissues (Fig. 4A). Mean adipocyte sizes in epididymal fat were greater in the three HFD-fed groups (especially in the HFD group) than in the ND group. However, the mean adipocyte size in the D-Kimchi group (1.8 ± 0.7 units) was smaller than that in the HFD or S-Kimchi groups (HFD group: 7.5 ± 2.6 units; S-Kimchi group: 3.4 ± 0.8 units), and was similar to that of the ND group (2.0 ± 0.7 units).
Furthermore, the two kimchi-treated groups had significantly lower MCP-1 mRNA expressions (an important mediator of macrophage activity and promoter of inflammatory response in adipose tissues) and IL-6 expressions (an inflammatory cytokine) in
epididymal fat tissues than the HFD group after the 8-week experimental period (Fig. 4B).
4. Discussion
In the present study, we investigated the antiobesity activities of home-made and commercial kimchi in Korea using a mouse model, and checked biomarkers in serum and tissue samples.
During the first 4 weeks of the experimental period, the weight gains shown by mice in the HFD, S-Kimchi, and D-Kimchi groups were similar. After the second 4 weeks, HFD group animals gained 13.5 g in body weight versus baseline, and the animals in the S-Kimchi, D-Kimchi, and ND group gained 3.7 g, 3.3 g, and 2.5 g, respectively. These figures are equal to body weight gains of only 0.1 ± 0.1 g/d in the S-Kimchi, D-Kimchi, and ND groups, but a
Fig. 4. Study of epididymal fat tissues of DIO mice (8th week). (A) Histological observations. (B) The mRNA expressions of MCP-1 and IL-6. The average sizes of adipocytes were measured on histological slides using Image J (right panel). The intensities of bands were measured by densitometry and are expressed as folds of the control (HFD) divided by GAPDH. Fold ratio: gene expression/GAPDH x control numerical value (control fold ratio set at 1). Group definitions are detailed in the legend of Table 1. *,t,z,x Means with different letters are significantly different (p < 0.05) by Duncan's multiple range tests. DIO, diet-induced obese; D-Kimchi, Korean commercial kimchi; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HFD, high-fat diet; IL-6, interleukin-6; MCP-1, monocyte chemotactic protein-1; ND, normal diet; S-Kimchi, standardized kimchi.
weight gain of 0.5 ± 0.1 g/d in the HFD group (Table 2). Furthermore, the mean food efficiency ratios in the S- and D-Kimchi groups (3.9 ± 0.1% and 3.5 ± 0.5%, respectively) and the ND group (2.7 ± 0.8%) were significantly lower than that in the HFD group (13.9 ± 1.7%) (p < 0.05), which confirms the body-weight-reducing effect of kimchi. In addition, kimchi (S- and D-Kimchi) significantly reduced adipose tissue weights (epididymal fat and perirenal fat) as compared with HFD alone (Fig. 1) (p < 0.05). The main subing-redients of kimchi, that is, red pepper powder, garlic, and ginger, have also been associated with significant reductions in body weight gain and fat mass in HFD-treated Sprague-Dawley (SD) rats [25]. S- and D-Kimchi contain similar levels of subingredients; however, D-Kimchi contains leeks and sea tangle, and is prepared using an Leu. mesenteroides DRC 0211 starter. Recently, it was reported that leeks have an antiobesity effect in vitro [26] and that Leuconostoc sp. JC046 decreases body weights of mice fed an HFD [27]. Thus, these subingredients might have affected the antiobe-sity effect observed in this study.
Long-term consumption of an HFD causes an imbalance in serum lipid profiles, as characterized by increased serum TG, TC, and LDL-c levels and reduced HDL-c levels, and this imbalance is a high risk factor of coronary heart disease and atherosclerosis [28]. In accordance with some other studies [26,29], we found that kimchi significantly reduced serum TG, TC, and LDL-c levels, and increased serum HDL-c levels in high-fat DIO mice. In addition, D-
Kimchi had significantly greater effects on LDL-c and HDL-c levels than S-Kimchi, which might have been associated with high levels of Leu. mesenteroides, which is in accord with the serum-lipid-controlling effect of Leuconostoc sp. in mice reported in another study [27]. Therefore, our results suggest that the bacterial composition in kimchi is related to the modulation of the serum lipid profile in DIO mice.
Adipose tissue releases adipocytokines, such as tumor necrosis factor alpha, IL-6, leptin, and adiponectin [30], and elevated serum leptin and depressed serum adiponectin levels are characteristic of obesity [31]. In another study, long-term intake of an HFD induced high levels of insulin in serum, caused insulin resistance, and increased fat accumulation in the liver [32]. In the present study, we found that kimchi decreased serum insulin and increased serum adiponectin, an insulin sensitizer that might improve insulin clearance and significantly reduce fat accumulation in the livers of DIO mice (Fig. 2). We also found that the two types of kimchi studied significantly reduced serum leptin levels and increased the levels of adiponectin, which are associated with adipocyte hypertrophy [33] and reduced fat accumulations in epididymal fat tissues (Figs. 2 and 4).
It is well known that the liver importantly regulates the accumulation of adipose tissue and controls lipid levels in the blood [34]. In previous studies, animals in the kimchi groups had lower liver weights (Fig. 1), and their livers contained smaller and fewer
lipid globules than animals in the HFD group (Fig. 3A). In particular, animals in the HFD group showed severe steatosis and hepatomegaly, but those in the kimchi groups had only moderate liver steatosis and showed no hepatomegaly. These results suggest that the administration of kimchi suppressed fat accumulation in the liver and, thus, reduced body weight gains.
SREBP-1c is an important lipogenic transcription factor that regulates the levels of fatty acid and TG by regulating the expression of FAS, a rate-limiting enzyme in the fatty acid pathway [35]. High levels of SREBP-1c induced by high HFD intake have also been reported to upregulate the expression of FAS, resulting in lipo-genesis in the liver and adipose tissue [35]. PPAR-g also enhances the activity of C/EBP, an important regulator of preadipocyte differentiation (C/EBP-b and C/EBP-5), and promotes adipogenesis (C/ EBP-a) [36]. In another study, reductions in PPAR-g activity effectively decreased body weight gains and hepatic TG levels in mice [37]. CPT-1 is a rate-limiting enzyme in b-oxidation of fatty acids and is used as a marker of lipid degradation [38]. In the present study, kimchi supplementation, particular D-Kimchi, significantly reduced hepatic mRNA levels of SREBP-1c, FAS, PPAR-g, and C/EBP-a, and increased CPT-1 mRNA levels, indicating that kimchi suppressed HFD-induced lipogenesis in DIO mice.
Obesity is associated with chronic systemic inflammation [39]. Elevated expressions of the mRNAs of tumor necrosis factor alpha, IL-6, and MCP-1 have been observed in the adipose tissues of obese mice [40], and in the present study, fat globules in epididymal fat tissues of the HFD group animals were larger than those of the ND group animals, whereas in kimchi supplement groups these were similar in size to that observed in the ND group (p < 0.05). This result suggests that kimchi reduced epididymal fat globule hypertrophy during the 4-week supplementation period. In addition, the S- and D-Kimchi groups showed significantly lower MCP-1 and IL-6 mRNA levels, and less macrophage infiltration into epididymal fat tissue than the HFD group (p < 0.05). These results suggest that kimchi has anti-inflammatory activity and positively reduces obesity in DIO mice.
Summarizing, the present study demonstrates the antiobesity effect of kimchi in a DIO mouse model. In this model, home-made and commercial kimchi suppressed body weight gains and fat accumulation in tissues, and modulated serum lipid levels and lipid metabolism-related genes in the liver and epididymal fat tissues. These results suggest the possibility that subingredients, such as leeks and starter (Leu. mesenteroides DRC 0211), enhance the anti-obesity effect of kimchi. Furthermore, our results show that commercial Korean kimchi can reduce body fat as well as home-made kimchi.
Conflicts of interest
All authors have no conflicts of interest to declare. Acknowledgments
This research was supported by grants (MCCM-B13006) from the Korean Heath Industry Development Institute (KHIDI) and by the National Center of Efficacy Evaluation for the Development of Health Products Targeting Digestive Disorders (NCEED).
References
[1] Park KY, Jeong JK, Lee YE and Daily 3rd JW. Health benefits of kimchi (Korean fermented vegetables) as a probiotic food. J Med Food 2014;17:6-20.
[2] Park KY. The nutritional evaluation and anti-mutagenic and anticancer effects of kimchi. J Kor Soc Food Sci Nutr 1995;24:169-82.
[3] Cho EJ, RHee SH, Lee SM and Park KY. In vitro antimutagenic and anticancer effects of kimchi fractions. J Kor Assoc Cancer Prev 1997;2:113-31.
[4] Lee YM, Kwon MJ, Kim JK, Suh HS, Choi JS and Song YO. Isolation and identification of active principle in Chinese cabbage kimchi responsible for antioxidant activity. Kor J Food Sci Technol 2004;36:129-33.
[5] Kim B, Park KY, Kim HY, Ahn SC and Cho EJ. Anti-aging effects and mechanisms of kimchi during fermentation under stress-induced premature senescence cellular system. Food Sci Biotechnol 2011;20:643-9.
[6] Noh JS, Kim HJ, Kwon MJ and Song YO. Active principle of kimchi, 3-(4'-hy-droxyl-3',5'-dimethoxyphenyl) propionic acid, retards fatty streak formation at aortic sinus of apolipoprotein E knockout mice. J Med Food 2009;12: 1206-12.
[7] Islam MS and Choi H. Antidiabetic effect of Korean traditional baechu (Chinese cabbage) kimchi in a type 2 diabetes model of rats. J Med Food 2009;12: 292-7.
[8] Jeong Y, Yang W, Kang J, Kong I and Kim J. Fibrinolysis of fermented kimchi. Kor J Life Sci 1995;5:203-10.
[9] Park JA, Tirupathi Pichiah PB, Yu JJ, Oh SH, Daily 3rd JW and Cha YS. Anti-obesity effect of kimchi fermented with Weissella koreensis OK1-6 as starter in high-fat diet-induced obese C57BL/6J mice. J Appl Microbiol 2012;113: 1507-16.
[10] Yoon JY, Jung KO, Kim SH and Park KY. Antiobesity effect of baek-kimchi (whitish baechu kimchi) in rats fed high fat diet. J Food Sci Nutr 2004;9: 259-64.
[11] Kim EK, An SY, Lee MS, Kim TH, Lee HK, Hwang WS, Choe SJ, Kim TY, Han SJ, Kim HJ, Kim DJ and Lee KW. Fermented kimchi reduces body weight and improves metabolic parameters in overweight and obese patients. Nutr Res 2011;31:436-43.
[12] Park KY and Rhee SH. Increased antimutagenic and anticancer activities of Chinese cabbage kimchi by changing kinds and levels of sub-ingredient. J Kor Soc Food Sci Nutr 1998;27:625-32.
[13] Choi SM, Kil JH, Jeon YS and Park KY. Fermentation characteristics, and anti-mutagenic and anticancer effects of mistletoe added kimchi. J Kor Assoc Cancer Prev 2003;8:98-106.
[14] Chang JY and Chang HC. Improvements in the quality and shelf life of kimchi by fermentation with the induced bacteriocin-producing strain, Leuconostoc citreum GJ7 as a starter. J Food Sci 2010;75:M103-10.
[15] Nonghyup Economic Research Institute. Korean kimchi industry trends and the research of consumer plan of Kimjang. 2013. p. 1. Report No.: 230.
[16] Li JJ, Huang CJ and Xie D. Anti-obesity effects of conjugated linoleic acid, docosahexaenoic acid, and eicosapentaenoic acid. Mol Nutr Food Res 2008;52: 631 -45.
[17] The Organisation for Economic Co-operation and Development [Internet]. OECD obesity update 2014. 2014 [cited 2015 Jun 25]. Available from: www. oecd.org/health/obesity-update.htm.
[18] Kersten S. Mechanisms of nutritional and hormonal regulation of lipogenesis. EMBO Rep 2001;2:282-6.
[19] Sung JH, Chon JW, Lee MA, Park JK, Woo JT and Park YK. The anti-obesity effect of Lethariella cladonioides in 3T3-L1 cells and obese mice. Nutr Res Pract 2011;5:503-10.
[20] Padwal RS and Majumdar SR. Drug treatments for obesity: orlistat, sibutr-amine, and rimonabant. Lancet 2007;369:71-7.
[21] Yun JW. Possible anti-obesity therapeutics from nature—a review. Phyto-chemistry 2010;71:1625-41.
[22] Saito M, Ueno M, Ogino S, Kubo K, Nagata J and Takeuchi M. High dose of Garcinia cambogia is effective in suppressing fat accumulation in developing male Zucker obese rats, but highly toxic to the testis. Food Chem Toxicol 2005;43:411-9.
[23] Jung KO, Kil JH, Kim KH and Park KY. Effect ofkimchi and its ingredients on the growth of Helicobacter pylori. Nutr Food 2003;8:149-53.
[24] Ministry of Food and Drug Safety (Korea). Production of food and food additives 2012. 2012. p. 58. Report No.: 14503.
[25] Yoon JY, Jung KO, Kil JH and Park KY. Antiobesity effect of major Korean spices (red pepper powder, garlic, and ginger) in rats fed high fat diet. J Food Sci Nutr 2005;10:58-63.
[26] Ahn EM, Kang HJ, Kim Y, Choe JS and Kang M. Effects of ethanol extracts from commonly consumed vegetables in Korea on differentiation and secretion of MCP-1 and adiponectin in 3T3-L1 adipocytes and lipid accumulation in HepG2 hepatocytes. J East Asian Soc Dietary Life 2015;25:99-110.
[27] Rico CW, Shin JH, Um IC and Kang MY. Cholesterol-lowering action and antioxidative effects of microbial gum in C57BL/6N mice fed a high fat diet. Biotech Bioprocess Eng 2011;16:167-72.
[28] Lavie CJ, Milani RV and Ventura HO. Obesity and cardiovascular disease: risk factor, paradox, and impact of weight loss. J Am Coll Cardiol 2009;53: 1925-32.
[29] Ku HS, Noh JS, Yun YR, Kim HJ, Kwon MJ, Cheigh HS and Song YO. Weight reduction and lipid lowering effects of sea tangle added Korean cabbage kimchi. J Kor Soc Food Sci Nutr 2007;36:1140-7.
[30] Lee WJ, Koh EH, Won JC, Kim MS, Park JY and Lee KU. Obesity: the role of hypothalamic AMP-activated protein kinase in body weight regulation. Int J Biochem Cell Biol 2005;37:2254-9.
[31] Lewis GF, Carpentier A, Adeli K and Giacca A. Disordered fat storage and mobilization in the pathogenesis of insulin resistance and type 2 diabetes. Endocr Rev 2002;23:201-29.
[32] Hao J, Liu SX, Zhao S, Liu QJ, Liu W and Duan HJ. High-fat diet causes increased serum insulin and glucose which synergistically lead to renal tubular lipid deposition and extracellular matrix accumulation. Br J Nutr 2012;107:74-85.
[33] Skurk T, Alberti-Huber C, Herder C and Hauner H. Relationship between adipocyte size and adipokine expression and secretion. J Clin Endocrinol Metab 2007;92:1023-33.
[34] Murase T, Nagasawa A, Suzuki J, Hase T and Tokimitsu I. Beneficial effects of tea catechins on diet-induced obesity: stimulation of lipid catabolism in the liver. Int J Obes Relat Metab Disord 2002;26:1459-64.
[35] Shimano H. SREBPs: physiology and pathophysiology of the SREBP family. FEBS J 2009;276:616-21.
[36] Farmer SR. Regulation of PPARg activity during adipogenesis. Int J Obesity 2005;29:S13-6.
[37] Jones JR, Barrick C, Kim KA, Lindner J, Blondeau B, Fujimoto Y, Shiota M, Kesterson RA, Kahn BB and Magnuson MA. Deletion of PPARg in adipose
tissues of mice protects against high fat diet-induced obesity and insulin resistance. Proc Natl Acad Sci U S A 2005;102:6207-12.
[38] Peng CH, Liu LK, Chuang CM, Chyau CC, Huang CN and Wang CJ. Mulberry water extracts possess an anti-obesity effect and ability to inhibit hepatic lipogenesis and promote lipolysis. J Agric Food Chem 2011;59: 2663-71.
[39] Wellen KE and Hotamisligil GS. Obesity-induced inflammatory changes in adipose tissue. J Clin Invest 2003;112:1785-8.
[40] Kang JH, Goto T, Han IS, Kawada T, Kim YM and Yu R. Dietary capsaicin reduces obesity induced insulin resistance and hepatic steatosis in obese mice fed a high-fat diet. Obesity 2010;18:780-7.
